FATTY-ACID BIOSYNTHESIS IN MYCOBACTERIUM-TUBERCULOSIS VAR BOVIS-BACILLUS-CALMETTE-GUERIN - PURIFICATION AND CHARACTERIZATION OF A NOVEL FATTY-ACID SYNTHASE, MYCOCEROSIC ACID SYNTHASE, WHICH ELONGATES N-FATTY ACYL-COA WITH METHYLMALONYL-COA

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  • 1 January 1985
    • journal article
    • research article
    • Vol. 260  (1) , 616-623
Abstract
A crude extract from Mycobacterium tuberculosis var. bovis BCG was previously shown to incorporate methylmalonyl-CoA into mycocerosic acids, exemplified by 2,4,6,8-tetramethyloctacosanic acid, and malonyl-CoA into n-fatty acids. The presence of several fatty acid synthases with differences in substrate preference and product chain length was detected in the crude extract of M. tuberculosis var. bovis. Among them was a mycocerosic acid synthase which was purified to homogeneity using anion-exchange chromatography, gel filtration, affinity chromatography and hydroxylapatite chromatography. This fatty acid synthase elongated long-chain fatty acyl-CoA primers using methylmalonyl-CoA and NADPH to produce multimethyl-branched mycocerosic acids. The enyzme was specific for methylmalonyl-CoA and would not incorporate malonyl-CoA into fatty acids. It elongated n-C6 to n-C20 CoA esters to generate primarily the corresponding tetramethyl-branched mycocerosic acids. Exogenous [1-14C]acyl-CoA and trideuteromethylmalonyl-CoA were incorporated into the multimethyl-branched fatty acids. Dodecyl sulfate electrophoresis showed that the enzyme had a MW of 238,000, whereas gel filtration showed a native MW of 490,000, indicating that the enzyme is composed of 2 monomers of identical MW. The enzyme contained on acyl carrier protein-like segment as indicated by incorporation of [I-14C] pantothenate into the 238-kDa [kilodaltons] protein and production of 1 mol of taurine/mol of the monomer upon hydrolysis of performic acid-oxidized enzyme. The mycocerosic acid synthase is a multifunctional enzyme similar to the well-characterized multifunctional fatty acid synthases except for the substrate specificity.

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